Utilizing Toxoplasma gondii To Understand Neuron-Specific Neuroinflammation

Toxoplasma gondii infects up to one third of the world’s population. The parasite can persist for the life of the host by hiding in neurons in the Central Nervous System (CNS). It was thought that neurons permitted this chronic infection due to a lack of innate immune capabilities including decreased expression of cytokine and immune cell receptors, as well as an inability to intracellularly clear the parasite. Evidence in the last five years has shown that neurons are capable of intracellular clearance, often in unusual ways, and are likely able to interact with cytotoxic immune cells. This brings into question the role of neurons during T. gondii infection as neurons are the primary CNS cell with which T. gondii interacts and persists in in vivo. The goal of this dissertation is to define T. gondii-neuron interaction in vivo. I first demonstrate that T. gondii’s neurotropism can be leveraged to deliver proteins to neurons. Then, I perform RNA-seq analysis to compare the expression of T. gondii-injected neurons (TINs) to neurons to neurons in an uninfected brain. These analyses revealed that TINs “transcriptomes” included immune cell transcripts, indicating the clustering of immune cells around TINs. By comparing this in vivo dataset with RNA-seq data from primary cortical neurons infected with either T. gondii or West Nile Virus, I determined that neurons that interact with T. gondii upregulate cytokine pathways, many of which are singular to T. gondii and not shared by virally infected neurons. Finally, to generate a better understanding of how T. gondii alters neurons in vivo, I have been pioneering single cell isolation and sequencing of TINs. This work has demonstrated that, in the infected brain, infiltrating immune cells must be removed in order isolate enough neurons for sequencing. In addition, neurons from infected brain are extremely fragile, suggesting that methods that do not require cell sorting and/or in which cells can first be fixed, will likely be necessary for this technique to be successful. Ultimately, the work in this dissertation highlights the complexity of neuron responses under different pathogens and paradigms and demonstrates the need for single cell analysis of neurons.

Name thesis:degree_name

Ph.D.

Level thesis:degree_level

doctoral

Discipline thesis:degree_discipline

Graduate College

Grantor dc:publisher

The University of Arizona.

Year dc:date.issued

2022

Author dc:creator

• Johnson, Hannah

Advisor dc:contributor.advisor

• Bhattacharya, Martha

Committee members dc:contributor.committeemember

• Rodgers, Kathleen
• Lybarger, Lonnie
• Koshy, Anita A.

• inflammation
• neuron
• RNA-seq
• Toxoplasma gondii
• transcriptomic

Statement dc:rights

• Copyright © is held by the author. Digital access to this material is made possible by the University Libraries, University of Arizona. Further transmission, reproduction, presentation (such as public display or performance) of protected items is prohibited except with permission of the author.

Licence dc:rights.uri

• http://rightsstatements.org/vocab/inC/1.0/

Language dc:language.iso

en

Handle dc:identifier.uri

http://hdl.handle.net/10150/668054

OAI identifier oai:identifier

oai:repository.arizona.edu:10150/668054